CN113736671A - Endophytic fungus AM8.2 for promoting absorption of potassium in oil tea - Google Patents
Endophytic fungus AM8.2 for promoting absorption of potassium in oil tea Download PDFInfo
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- CN113736671A CN113736671A CN202111222600.9A CN202111222600A CN113736671A CN 113736671 A CN113736671 A CN 113736671A CN 202111222600 A CN202111222600 A CN 202111222600A CN 113736671 A CN113736671 A CN 113736671A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
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- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
- C05F11/00—Other organic fertilisers
- C05F11/08—Organic fertilisers containing added bacterial cultures, mycelia or the like
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- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
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- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
- C05G3/80—Soil conditioners
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W30/00—Technologies for solid waste management
- Y02W30/40—Bio-organic fraction processing; Production of fertilisers from the organic fraction of waste or refuse
Abstract
The invention provides endophytic fungus AM8.2 for promoting potassium element absorption of oil tea, which is obtained by screening roots of woody plant oil tea, and belongs to the technical field of microorganisms. The classification name of the endophytic fungus AM8.2 is acanthrax anthrax (Colletotrichum spinosum.) AM8.2, which was deposited in China general microorganisms of the Committee for culture Collection of microorganisms at 09 month 01, 2021The preservation number of the core is CGMCC No. 23245. The bacterial manure prepared from the endophytic fungi AM8.2 can reduce the use of chemical fertilizers, improve the microbial activity of soil and influence the physical and chemical properties of the soil of the oil tea, thereby influencing the absorption of endogenous hormones, potassium and nitrogen elements of the oil tea and promoting the growth and development of plants.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to endophytic fungus AM8.2 for promoting potassium element absorption of oil tea.
Background
Potassium is an essential element for the growth and development of crops, and the potassium content in the plant bodies generally accounts for 0.2-4.1 percent of the dry matter and is only lower than that of nitrogen. It can regulate plant metabolism process, promote carbohydrate synthesis and nitrogen metabolism, raise crop yield, improve fruit quality and strengthen plant leaf photosynthesis. Can also promote the transportation of the photosynthetic products to tubers, and can improve the synthesis capacity of sucrose in leaves. The potassium can enhance the stress resistance of crops and promote the formation of good quality of the crops.
Oil tea (tea-oil camellia)Camellia oleifera Abel.) Belonging to the genus Camellia of the family Theaceae, together with oil palm, olive and coconut, are called the world's four woody oil tree species. The camellia oleifera is an important woody edible oil crop in hilly areas in south China, has more than 1160 billion annual output values, and plays a very important role in forestry industry and economic development of hilly and mountain areas. Meanwhile, the camellia oleifera leaves can adsorb harmful substances in the air, and have certain ecological value. From the actual planting condition of the camellia oleifera in China, the yield of most camellia oleifera in China is not high, and the production enthusiasm of growers is greatly struck. The low yield and low yield are the main problems facing the oil tea industry at present, and the healthy and sustainable development of the oil tea industry can be promoted only by improving the yield of the oil tea. Mineral nutrient element for camellia oleiferaThe growth and yield of the camellia oleifera are obviously influenced, nearly one fourth of land is lack of potassium in China, the operator has insufficient attention on the potassium fertilizer, potassium becomes a main factor influencing the growth and quality of crops, and the potassium absorption rate of the camellia oleifera is required to be improved for promoting the growth and yield increase of the camellia oleifera.
Scholars at home and abroad screen various endophytic fungi with growth promotion from plants, but the current research mainly focuses on screening related strains of gramineae plants, and the research on screening and identifying woody plants, particularly the endophytic fungi of camellia oleifera, is rarely reported. The endophytic fungi beneficial to the growth and stress resistance of the oil-tea camellia is screened from the oil-tea camellia root, an oil-tea camellia-endophytic fungi symbiotic system is established, so that the endophytic fungi capable of promoting the growth of the oil-tea camellia and improving the potassium absorption of the oil-tea camellia are obtained, and a data basis and a reference basis are provided for enriching strain information of the endophytic fungi of woody plants and producing biological bacterial fertilizers for the oil-tea camellia forest management.
Disclosure of Invention
The invention aims to provide an endophytic fungus AM8.2 for promoting potassium element absorption of camellia oleifera aiming at the problem that the woody plant camellia oleifera lacks a camellia oleifera-endophytic fungus symbiotic system.
In order to achieve the purpose, the invention adopts the following technical scheme:
an endophytic fungus AM8.2 for promoting the absorption of potassium element in oil tea, the classification of the endophytic fungus AM8.2 is named as acanthrocarpus (A) (B)Colletotrichum spinosum.) AM8.2, which has been deposited in China general microbiological culture Collection center on the 09.01.2021 year old, with the collection number of CGMCC No.23245 and the collection address of No. 3, Siro No.1, Beijing, the area facing the Yangyang province.
The application of the endophytic fungus AM8.2 for promoting the absorption of potassium element in the camellia oleifera in promoting the growth of the camellia oleifera is provided.
The endophytic fungus AM8.2 is obtained by separating and purifying from the roots of the camellia oleifera, can be prepared into bacterial liquid, and can be prepared into bacterial liquid for promoting the growth of camellia oleifera seedlings by mixing with organic fertilizers and applying in furrow.
A bacterial fertilizer containing endophytic fungi AM8.2 bacterial liquid comprises the following components: the endophytic fungi AM8.2 bacterial liquid and organic fertilizer are mixed according to the proportion of 1 liter: 10 kg ofThe components are mixed according to the proportion. The preparation method of the endophytic fungus AM8.2 bacterial liquid comprises the following steps: inoculating endophytic fungus AM8.2 into liquid culture medium, culturing at 28 deg.C for 72 hr in 220 r/min shaking table, and diluting the obtained culture solution with sterile water to spore concentration of 0.6 × 109CFU/ml. The formula of the liquid culture medium is as follows: 20.0g of peptone, 10.0g of yeast extract, 20.0g of glucose, 0.03g of adenine sulfate and 1000ml of ultrapure water, wherein the pH value is 5.6 +/-0.2.
The bacterial fertilizer containing the endophytic fungus AM8.2 bacterial liquid is applied to planting of the camellia oleifera.
The invention has the advantages that:
the endophytic fungus AM8.2 is obtained by screening roots of woody plant oil tea, and can promote the oil tea to absorb potassium elements and nitrogen elements, so that the plant growth can be promoted. The bacterial manure prepared from the endophytic fungus AM8.2 can reduce the use of chemical fertilizers, improve the microbial activity of soil and influence the physical and chemical properties of the soil of the oil tea, thereby influencing the absorption of endogenous hormones, potassium and nitrogen elements of the oil tea. Thereby promoting the growth and development of plants.
Description of the drawings:
FIG. 1 is a colony morphology diagram of endophytic fungus AM 8.2. Left: a front side; and (3) right: and a back surface.
Detailed Description
In order to make the present invention more comprehensible, the technical solutions of the present invention are further described below with reference to specific embodiments, but the present invention is not limited thereto.
Example 1 isolation of Camellia oleifera root endophytic fungi
Culture medium:
(1) liquid medium YPDA: peptone 20.0g, yeast extract 10.0g, glucose (C)6H12O6•H2O) 20.0g, adenine sulfate 0.03g, ultra pure water 1000ml, pH 5.6 + -0.2.
(2) PDA medium (potato dextrose agar medium): 6.0g of potato extract powder, 20.0g of agar, 20.0g of glucose and 5.6 +/-0.2 of pH.
Isolation of endophytic fungi
(1) Adopting the tissue separation method, after the root of the oil tea is washed clean by running water, drying the oil tea in a super clean bench for tissue surface disinfection, wherein the operation flow is as follows: 75vol% absolute ethyl alcohol sterilization → sterile water cleaning 2-3 times → 15vol% sodium hypochlorite sterilization → sterile water cleaning 2-3 times. Cutting off phloem of the sterilized root with sterile blade, cutting into size of 2mm × 2mm, placing on PDA culture medium, and culturing at 28 deg.C in dark place.
(2) And (3) verification of the disinfection effect: and (3) coating sterile water for cleaning the sample in the last step of disinfection on an unused PDA culture medium, and culturing at a constant temperature of 28 ℃ for 4-7 days, wherein if no thallus grows out, the sample is disinfected completely. And (3) adopting a tissue blotting method, slightly rolling the sterilized sample tissue on an unused PDA culture medium or tightly adhering to the culture medium for 5min, taking away the sample tissue for comparison, and culturing at the constant temperature of 28 ℃ for 4-7 d, wherein the sample tissue is sterilized if no thallus grows out. Each control was repeated 3 times.
Purification of endophytic fungi
After the tissue material is cultured for 3-5 days, hyphae with good growth of bacterial colonies around the tissue are picked by an inoculating needle, the hyphae are respectively purified on a PDA culture medium by adopting a scribing method, and are inversely placed in a constant-temperature incubator and are cultured for 4-7 days at a constant temperature and in a dark place. And repeatedly purifying for 3-4 times to obtain a purified strain AM 8.2. Inoculating the purified strain into PDA slant culture medium, and storing at 4 deg.C.
3 morphological characteristics of endophytic fungus AM 8.2:
4 DNA identification of endophytic fungi
Amplifying an 18S rDNA gene sequence (SEQ ID NO. 1) of the endophytic fungus AM8.2 by using fungus 18S rDNA universal forward and reverse primers (ITS 1: TCCGTAGGTGAACCTGCGG, ITS 4: TCCTCCGCTTATTGATATGC); the result of comparing the 18S rDNA sequence with the sequence in GenBank database shows that AM8.2 strain and anthrax acantha (B)Colletotrichum spinosum.) On one branch; combined with colony morphology, physiological and biochemical characteristics and 18S rDNA sequence analysis, the primary identification is acantha anthrax (Colletotrichum spinosum.）。
>AM8.2 SEQ ID NO.1：
TCTTCCGTAAGGGTGACCTGCGGAGGGATCATTACTGAGTTTACGCTCTATAACCCTTTGTGAACATACCTAACCGTTGCTTCGGCGGGCGGGAGGTCCGCCTCCCCCCCGGCCCCGCTCGCGGGGCGCCCGCCGGAGGAAAAACCCAACTCTTATTTTAACGACGTCTCTTCTGAGTGGCACAAGCAAATAATCAAAACTTTTAACAACGGATCTCTTGGTTCTGGCATCGATGAAGAACGCAGCGAAATGCGATAAGTAATGTGAATTGCAGAATTCAGTGAATCATCGAATCTTTGAACGCACATTGCGCCCGCCAGCATTCTGGCGGGCATGCCTGTTCGAGCGTCATTTCAACCCTCAAGCACCGCTTGGCGTTGGGGCTTCCACGGCTGACGTGGGCCCTCAAAGACAGTGGCGGACCCTCGCGGAGCCTCCTTTGCGTAGTAACATACCACCTCGCACCGGGACCCGCAGGGCACTCCTGCCGTAAAACCCCCCAATTTTTACAAGGTTGACCTCGGATCAGGTAGGAATACCCGCTGAACTTAAGCATATCTAAAGCCGGAAGAAAAAATAAC
Example 2
Preparing bacterial liquid: inoculating activated endophytic fungus AM8.2 into 20mL potato glucose liquid culture medium, culturing on constant temperature shaking table for 72h (28 deg.C, 220 r/min), sucking 1mL, inoculating into 2 500mL LYPDA culture medium, counting spore number with blood counting plate, diluting with sterile water to spore concentration of 0.6 × 109CFU/ml。
Preparing bacterial manure: the mixed solid bacterial fertilizer comprises the following components: endophytic fungus AM8.2 bacterial liquid: organic fertilizer = 1 liter: 10 kg. The organic fertilizer is prepared by fermenting tobacco powder, rice bran, cow dung, peanut residue, plant ash, humic acid, sheep dung and the like, and the mass fraction of organic matters is more than 46 percent, and the mass fraction of N + P2O5+K2The mass fraction of O is more than 5 percent.
The experimental scheme is as follows: the test adopts an outdoor test, and the camellia oleifera forest field selected in the test is provided by the available forest field of Qingliu county, Qingliu city, Sanming City, Fujian province. And selecting the oil-tea camellia forest with good growth state as a sampling area in the bacterial manure experiment area and 1 blank control area. The prepared solid bacterial manure is applied to a bacterial manure test area in a watering mode, and is applied in a furrow mode at a position 40 centimeters below a projection area of a camellia oleifera forest land in 1 month, and soil samples and leaf samples are collected at the bottom of 6 months because 7 months are rainy seasons. And randomly selecting three sampling points to sample leaves, roots and soil, and mixing 5 parts of samples into 1 sample to be detected at each sampling point.
Influence of bacterial manure on soil physicochemical properties
Three physical and chemical properties such as water content, conductivity and pH value of the soil sample are measured, and the results are shown in Table 1:
TABLE 1 soil moisture content, conductivity and pH
Different lower case letters indicate significant differences at the same treatment level (t test,p < 0.05）
as can be seen from table 1, the water content, conductivity and PH of the bacterial manure prepared from the AM8.2 strain were significantly different from those of the control group.
Influence of bacterial manure on endogenous hormones in camellia oleifera leaves
The concentrations of the standards in the Plant GA ELISA KIT (ml 072782, MLBIO, Shanghai), Plant IAA ELISA KIT (ml 022829, MLBIO, Shanghai), Plant ABA ELISA KIT (ml 064270, MLBIO, Shanghai) and Plant BR ELISA KIT (ml 036309, MLBIO, Shanghai) KITs were used as abscissa, and the measured OD values were used as ordinate, and a standard curve was drawn to obtain a linear regression equation, and the sample content was calculated (Table 2).
TABLE 2 GA, IAA, ABA, BR content
Different lower case letters indicate significant differences at the same treatment level (t test,p < 0.05）
as can be seen from Table 2, the bacterial manure prepared from the AM8.2 strain had GA (pmol/ml) content of 211.30. + -. 11.92 and ABA (ng/ml) content of 140.26. + -. 4.51, which was significantly different from the control group, as compared with the control group.
Influence of bacterial manure on absorption of nitrogen, phosphorus and potassium elements by camellia oleifera
The relative contents of N, P, K and other three elements endogenous in the camellia oleifera leaves are measured, and the results are shown in table 3:
TABLE 3 relative contents of three elements, endogenous N, P, K, etc
Different lower case letters indicate significant differences at the same treatment level (t test,p < 0.05）
as can be seen from table 3, when compared with the control treatment group, the bacterial manure prepared from the AM8.2 strain had a relative content of K of 1.39 ± 0.10, which is significantly different from the control group, indicating that the strain AM8.2 can significantly promote the absorption of potassium (K) by camellia oleifera; meanwhile, the relative content of N is 1.56 +/-0.21, and is also obviously improved compared with a control group.
The soil is a necessary condition for plant growth and development and nutrient substance absorption, the physical and chemical properties of the soil are comprehensively reflected by climate, vegetation, terrain and soil structure, and the application of microbial fertilizer can reduce the use of chemical fertilizer and improve the microbial activity of the soil, thereby influencing the growth and development of plants. From the above results, the experiment can obtain that the organic bacterial manure number prepared from the endophytic fungus AM8.2 has positive influence on the soil growth environment of the oil tea, and can influence the soil physicochemical properties of the oil tea to a certain extent, so that the absorption of endogenous hormones and potassium of the oil tea is influenced.
The above description is only a preferred embodiment of the present invention, and all equivalent changes and modifications made in accordance with the claims of the present invention should be covered by the present invention.
SEQUENCE LISTING
<110> Wangxiao shou, Wuxuanyi and Wangxiao Wei
<120> endophytic fungus AM8.2 for promoting absorption of potassium in oil-tea camellia
<130> 3
<160> 3
<170> PatentIn version 3.3
<210> 1
<211> 581
<212> DNA
<213> SEQ ID NO.1
<400> 1
tcttccgtaa gggtgacctg cggagggatc attactgagt ttacgctcta taaccctttg 60
tgaacatacc taaccgttgc ttcggcgggc gggaggtccg cctccccccc ggccccgctc 120
gcggggcgcc cgccggagga aaaacccaac tcttatttta acgacgtctc ttctgagtgg 180
cacaagcaaa taatcaaaac ttttaacaac ggatctcttg gttctggcat cgatgaagaa 240
cgcagcgaaa tgcgataagt aatgtgaatt gcagaattca gtgaatcatc gaatctttga 300
acgcacattg cgcccgccag cattctggcg ggcatgcctg ttcgagcgtc atttcaaccc 360
tcaagcaccg cttggcgttg gggcttccac ggctgacgtg ggccctcaaa gacagtggcg 420
gaccctcgcg gagcctcctt tgcgtagtaa cataccacct cgcaccggga cccgcagggc 480
actcctgccg taaaaccccc caatttttac aaggttgacc tcggatcagg taggaatacc 540
cgctgaactt aagcatatct aaagccggaa gaaaaaataa c 581
<210> 2
<211> 19
<212> DNA
<213> ITS1
<400> 2
tccgtaggtg aacctgcgg 19
<210> 3
<211> 20
<212> DNA
<213> ITS4
<400> 3
tcctccgctt attgatatgc 20
Claims (5)
1. An endophytic fungus AM8.2 for promoting the absorption of potassium element in oil tea, which is characterized in that: the classification name of the endophytic fungus AM8.2 is acanthrax anthrax (Colletotrichum spinosum.) AM8.2, which has been deposited in China general microbiological culture Collection center on the 09.01.2021 year old, with the collection number of CGMCC No.23245 and the collection address of No. 3, Siro No.1, Beijing, the area facing the Yangyang province.
2. The application of the endophytic fungus AM8.2 for promoting the absorption of potassium element in the camellia oleifera as claimed in claim 1 in promoting the growth of the camellia oleifera.
3. A bacterial fertilizer containing endophytic fungi AM8.2 bacterial liquid is characterized in that: the preparation of the bacterial manure comprises the following steps: the endophytic fungi AM8.2 bacterial liquid and organic fertilizer are mixed according to the proportion of 1 liter: mixing 10 kg of the mixture into solid bacterial manure; the preparation method of the endophytic fungus AM8.2 bacterial liquid comprises the following steps: inoculating endophytic fungus AM8.2 into liquid culture medium, culturing at 28 deg.C for 72 hr in 220 r/min shaking table, and diluting the obtained culture solution with sterile water to spore concentration of 0.6 × 109 CFU/ml。
4. A bacterial manure according to claim 3, characterized in that: the formula of the liquid culture medium is as follows: 20.0g of peptone, 10.0g of yeast extract, 20.0g of glucose, 0.03g of adenine sulfate and 1000ml of ultrapure water, wherein the pH value is 5.6 +/-0.2.
5. The use of the bacterial manure of claim 3 in camellia oleifera plantation.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114921362A (en) * | 2022-04-19 | 2022-08-19 | 湖南大学 | Camellia oleifera endophytic actinomycetes with insoluble phosphorus dissolving and growth promoting functions and application thereof |
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| CN102132672A (en) * | 2010-12-09 | 2011-07-27 | 海南珠峰园林科技有限公司 | Method for cultivating epiphytic orchid |
| CN105132332A (en) * | 2015-09-28 | 2015-12-09 | 中国科学院天津工业生物技术研究所 | Acetobacter gluconicum and application of acetobacter gluconicum as plant growth-promoting rhizobacteria |
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| CN101338290A (en) * | 2008-08-12 | 2009-01-07 | 金辉 | Method for culturing orchid special strain thereof |
| CN102132672A (en) * | 2010-12-09 | 2011-07-27 | 海南珠峰园林科技有限公司 | Method for cultivating epiphytic orchid |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN114921362A (en) * | 2022-04-19 | 2022-08-19 | 湖南大学 | Camellia oleifera endophytic actinomycetes with insoluble phosphorus dissolving and growth promoting functions and application thereof |
| CN114921362B (en) * | 2022-04-19 | 2023-07-25 | 湖南大学 | Oil tea endophyte with functions of dissolving insoluble phosphorus and promoting growth and application thereof |
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